A reliable skin test antigen for use in the diagnosis and epideminological investigation of blastomycosis is not presently available. Previous approaches to this problem have, for the most part, been limited to analyses of crude, broth culture filtrates of mycelial or yeast cells. In an effort to elucidate the component responsible for the delayed hypersensitivity response, cell walls and cytoplasmic components will be isolated from mechanically disrupted yeast-phase cells of Blastomyces dermatitidis. Alkaline hydrolysis, mild acid hydrolysis, and phenol will be employed to solubilize yeast-phase wall constituents. The cell wall fractions and soluble cytoplasmic components will be partially purified by column chromatography and ultrafiltration. For skin test studies, each fraction will be inoculated intradermally into Blastomyces-infected guinea pigs. To evaluate the specificity of any fraction(s) eliciting a dermal hypersensitivity response in homologously infected animals, groups of unifected, histoplasma-infected, and Coccidioides-infected guinea pigs will be skin tested. The efficacy of the antigen(s) will be further assessed in the in vitro macrophage inhibition assay using peritoneal exudate cells obtained from each guinea pig group. Chemical analyses will be employed to determine the nature of the skin-test active component. In preliminary analyses, acid hydrolyzed fractions will be analyzed for amino acids, carbohydrate, lipid, and nucleic acid content. Conventional methods will then be employed to determine the molecular weight, electrophoretic properties, and homogeniety of the yeast-phase antigen.